Cloning And Functional Analyses Of AmDREB1 And AmDREB2 Genes From Ammopiptanthus Mongloicus

DREB(Dehydration responsive element binding) transcription factors play important roles in the regulatory network of stress-resistant gene expression in plants. At present, DREBs are one of the most focused regulatory factors in the studies on molecular mechanism and gene engineering of plant stress resistance. In this paper, both cDNA and genomic DNA fragments of AmDREB1 were cloned from Ammopiptanthus mongolicus, a strong stress-resistant plant, by PCR method. Moreover, the expression patterns and functions of AmDREB1 and AmDREB2 which was cloned previously in our laboratory were analyzed. The main results are as follows:1. The expression analysis of AmDREB2 by semi-quantitative RT-PCR indicated that this gene was involved in the response of A.mongolicus to exogenous ABA.2. The cDNA fragment of AmDREB2 was successfully ligased to plant over-expression vector pCAMBIA3301 and then was introduced into Arabidopsis by Agrobacterium-mediated transformation. The transgenic plants were screened and identified, and the homozygous lines of T3 generation were obtained. Overexpression of AmDREB2 in Arabidopsis did not show growth inhibition and enhanced the resistance of the transgenic plants to salt and drought stresses.3. In the AmDREB2 transgenic lines, the expression levels of RAB18 and RD29 A, two stress marker genes, and the Na+/H+ antiporter gene AtNHX1 were higher than those in wild type plants, indicating that these genes were regulated by the transgenic gene AmDREB2.4. The complete coding region cDNA and genomic DNA of AmDREB1 were cloned by PCR method. Both the regions consist of 474 bp, indicating no introns in this gene. The predicted protein of AmDREB1 consists of 158 amino acid residues, with a molecular weight of 39.1 kDa and an isoelectric point of 5.2. The protein was predicted to contain an AP2 domain and a nuclear localization signal and most likely locates in nucleus. Semi-quantitative RT-PCR analysis showed that the expression of AmDREB1 was induced by high salinity, exogenous ABA, drought and cold. Specifically, the induction was stronger under cold and drought stresses than in other two conditions. The cDNA fragment of AmDREB1 was inserted into plant inducible expression vector and was further transformed into Arabidopsis. Through preliminary identification, of the transgenic plants, it was found that AmDREB1 could not markedly enhance the tolerance to salt stress.